Antimicrobial peptides as models for the evolution of gene duplication

 描述（由申请人提供）：作为基因复制进化模型的抗菌肽抗菌肽（AMP）是真核生物体液免疫应答的重要组成部分。通过复制的基因家族扩展长期以来被认为是产生进化新颖性的一种手段，尽管从基因复制到新功能的进化过程还没有很好地建立。进化的一个显著特征是 AMP基因家族中的高重复率是AMP基因家族中的一个重要特征。这项研究将使用AMP基因家族中的重复，以更好地了解AMP功能和基因重复的进化。使用AMP研究基因复制可能适用于人类健康，并提供了果蝇家族中基因复制的内置复制。此外，在果蝇中进行高通量实验和遗传操作的能力以及一组参与AMP功能的预期表型的存在使得该系统易于处理。目的1是一个全面的分析AMP的进化内和物种之间，以解决的作用，选择在进化中发挥基因复制后，以及基因表达的分歧与物种和旁系同源。该分析的一部分将在空间（跨组织）和时间上以非常精细的尺度检查这些AMP的表达。目标1的训练部分包括最先进的群体遗传分析以及精细规模的基因表达研究。目的2包括使用工程基因敲除并采用大量病原体来解决特异性AMP旁系同源物在响应口腔感染和全身感染中的功能，因为我们有证据表明AMP免疫应答是病原体特异性的。根据本实验的结果，我们将创建具有多个拷贝的特定AMP的转基因系，以测试基因剂量的影响，在旁系同源物之间进行启动子交换实验，以测试调节序列的变化如何影响亚功能化过程中的表达。目标2的培训包括使用开发中的基因组编辑技术构建转基因品系，以解决这些问题。目标3将特别关注AMP重复在自然群体中的分离，以解决最初重复后不久的parabolic功能的性质 活动我们将采用与目标1和2相同的基本框架，检查变异性、基因表达和parabolic功能的模式，但侧重于这些分离的串联重复。我们还将创建转基因串联重复，以进行控制背景效应的功能测定。由于AMP基因重复在基因家族中的复制性质，我们期望得出关于基因重复进化的结论。拟议研究的培训部分将补充申请人以前的经验，并将他定位为富有成效的研究生涯。康奈尔大学和拉扎罗和克拉克实验室共同拥有资源和专业知识，以确保成功完成赠款的培训阶段。

 DESCRIPTION (provided by applicant): Antimicrobial peptides as models for the evolution of gene duplication Antimicrobial peptides (AMPs) are a vital part of the humoral immune response for eukaryotes. Gene family expansion through duplication has long been recognized as a means of generating evolutionary novelty though the evolutionary processes leading from gene duplication to novel function is not well established. One striking characteristic of the evolution of AMPs is the high rate of gene duplication in AMP gene families. The proposed research will use duplications in AMP gene families to better understand both AMP function and the evolution of gene duplication. Using AMPs to study gene duplication may have applicability to human health and provides built-in replication of gene duplications across families in Drosophila. Furthermore, the ability to perform high throughput experiments and genetic manipulations in Drosophila and the existence of a set of expected phenotypes involved in AMP function make the system tractable. Aim 1 is a comprehensive analysis of AMP evolution within and between species to address the role that selection plays in evolution after gene duplication and how gene expression diverges with between species and paralogs. Part of this analysis will examine expression of these AMPs on a very fine scale both spatially (across tissues) and temporally. The training component of Aim 1 includes state-of-the art population genetic analyses as well as fine scale gene expression studies. Aim 2 consists addresses the function of specific AMP paralogs in response to both oral infection and systemic infection using engineered gene knockouts and employing a large variety of pathogens since we have evidence that AMP immune response is pathogen-specific. Based on the results from this experiment, we will create transgenic lines with multiple copies of a particular AMP to test the effects of gene dosage, perform promoter swapping experiments between paralogs to test how changes in regulatory sequence might influence expression en route to subfunctionalization. Training in Aim 2 consists of the construction of transgenic lines using developing genome editing technologies to address these questions. Aim 3 will focus specifically on AMP duplicates segregating in natural populations to address the nature of paralog function shortly after the initial duplication event. We will employ the same basic framework as in Aims 1 and 2 examining patterns of variability, gene expression and paralog function but focusing on these segregating tandem duplicates. We will additionally create transgenic tandem duplicates to perform functional assays controlling for background effects. Because of the replicated nature of AMP gene duplicates across gene families, we expect to draw conclusions about the evolution of gene duplication. The training portion of the proposed research will complement the applicant's previous experience and position him for a productive research career. Cornell University and the Lazzaro and Clark labs together have the resources and expertise to ensure the successful completion of the training phase of the grant.